We request funds to purchase and install an NMR spectrometer system for highly sensitive direct detection of signals from carbon-13 and nitrogen-15 in proteins, nucleic acids, metabolites, and natural products. The system will have multinuclear (hydrogen-1, hydrogen-2, carbon-13, and nitrogen-15) capability. To save money, this system will re-use an Oxford 500 MHz (1H) magnet available in house. Direct observation of carbon-13 and nitrogen-15 has proven to be the only way to detect NMR signals from the active sites of paramagnetic metal containing proteins. In addition, direct observation of carbon-13 has shown great promise as a strategy for collecting and assigning NMR spectral data from larger proteins and nucleic acids. The user projects described cover structure-function investigations of proteins, protein-protein complexes, protein-ligand complexes, nucleic acids, protein-nucleic acid complexes, and natural products. The systems to be studied are large, complex, and in several cases contain paramagnetic centers. These properties preclude or hinder their investigation by standard proton-detected multinuclear approaches. We provide examples that demonstrate the necessity of the direct detection approach. Compared to currently available instrumentation, the requested system will increase the sensitivity of direct carbon-13 and nitrogen-15 detection experiments by a factor of 3-4 and thus decrease data collection time by a factor of 10. The system will enable the investigation of systems that are larger, less-soluble, and less-stable than can be currently considered. The instrumentation will be installed in the National Magnetic Resonance Facility at Madison (NMRFAM) in space already fitted out and optimized for tight control of temperature and humidity and with conditioned power and a captive chilled-water system. The instrumentation will be managed by spectroscopists experienced in the operation and maintenance of NMR systems with cryogenic probes, who will provide training in the use of the instrumentation and work with users to develop optimal data collection approaches specific to their problems. Access will be allocated according to published NMRFAM operating procedures, and oversight will be provided by NMRFAM's standing local and external advisory boards. Direct-detection experiments offer one the most exciting new approaches to structure-function investigations of proteins and nucleic acids. Among the systems described are ones of direct relevance to human disease: iron-sulfur assembly (Friedreich's ataxia), fatty acid desaturation (obesity, diabetes), and RNA-metal interactions (ageing, cancer). [unreadable] [unreadable] [unreadable]